HIF-1 controls the transcription of many genes that are involved in key aspects of cancer biology. Overexpression of HIF-11, an inducible subunit of HIF-1 in response to hypoxia, is a prognostic factor in many cancers. The major signaling axis mediated by PI3K, PDK1, and Akt (also referred as protein kinase B) plays a significant role in the regulation of HIF-11 expression. Both GSK32 and MAPKs are also known to directly phosphorylate HIF-11, thereby affecting its stability and/or nuclear localization. To understand the regulatory network that suppresses tumor development and tumor angiogenesis, we have recently identified HIF-11 as a new in vitro substrate of Polo-like kinase 3 (Plk3). Plk3 strongly phosphorylates HIF-11 on serines 576 and 657 in vitro, two residues that lie in the oxygen-dependent degradation domain and near the nuclear export signal, respectively. By studying primary isogenic murine embryonic fibroblasts (MEFs), we have shown that PLK3-/- MEFs are hyper-sensitive to the induction of HIF-11 under hypoxia or treated with nickel, a hypoxia mimetic. Compared with that of wild- type MEFs, PLK3-/- MEFs contain a high level of Akt1/PKB activities, which is tightly associated with the inhibitory phosphorylation of GSK32. Consistent with the potential role of Plk3 in regulating the hypoxia signaling network, PLK3-/- mice develop tumors in various organs at an advanced age and PLK3-/- tumors are large in size and highly vascularized, suggesting active tumor angiogenesis. On the basis of these observations regarding physical and functional interactions between Plk3 and HIF-11, we hypothesize that Plk3 negatively regulates the hypoxia regulatory network and HIF-11-dependent tumor angiogenesis. To test the validity of this hypothesis, we will (i) study functional interaction between Plk3 and known signaling molecules, including GSK32 and MAPKs that phosphorylate HIF-11, and identify additional Plk3 target(s) upstream of Akt1, (ii) determine whether PLK3-/- mice are prone to tumorigenesis under hypoxia, and (iii) investigate whether mice harboring Plk3 phosphorylation-resistant mutant alleles of HIF-11 are more susceptible to tumorigenesis after nickel exposure. The combined in vitro and in vivo studies will greatly facilitate the elucidation of a new mechanism by which HIF-11 is regulated by Plk3 during hypoxic responses or after exposure to environmental carcinogens such as nickel compounds. A detailed understanding of the molecular regulation of HIF-11 will add significantly to the existing knowledge of tumor angiogenesis and tumor cell resistance to anti-cancer therapies.